Ac-SDKP may mediate some of the anti-fibrotic effects of ACE inhibitors. However, very little is known about the in vivo synthesis of this peptide, save that thymosin-beta4 is the most likely precursor. We recently demonstrated that the enzyme responsible for release of the anti-fibrotic peptide from its substrate is prolyl oligopeptidase (POP); however, this enzyme cleaves peptides no larger than 30 amino acids, so that some other peptidase, possibly chymase, may shorten the substrate prior to POP hydrolysis. We propose to elucidate a) the enzymes involved in the production of Ac-SDKP; b) the physiological role of endogenous Ac- SDKP in preventing fibrosis and inflammation during ACEi treatment; and c) whether or not expression and/or activity of the enzyme responsible for peptide synthesis are modified in pathological conditions such as hypertension or during treatment with ACEi. In aim 1 we hypothesize that chymase is involved in the enzymatic cleavage of thymosin-beta4 prior to POP hydrolysis and release of Ac-SDKP. We propose to study: 1) in vitro production of Ac-SDKP from thymosin-beta4 by heart, kidney, blood vessel and leukocyte homogenates in the presence or absence of chymase inhibitors; and 2) in vivo production of Ac-SDKP after treatment with chymase and/or ACE inhibitors. In aim 2 we hypothesize that prolyl isomerase participates in the production of Ac-SDKP from thymosin-beta4, converting the substrate at its trans conformati on prior to POP hydrolysis. We propose to study: 1) in vitro production of Ac-SDKP from thymosin-beta4 in the presence or absence of prolyl isomerase inhibitors; and 2) in vivo production of Ac-SDKP in different tissues after treatment with prolyl isomerase inhibitors and/or ACE inhibitors. In specific aims 1 and 2 we will clarify the enzymes involved in the cleavage of thymosin-beta4 to release Ac-SDKP. In aim 3 we hypothesize that endogenous Ac-SDKP plays a physiological role in antagonizing the effects of pro-fibrotic and proinflammatory stimuli such as angiotensin II (Ang II). We propose to study whether a specific POP inhibitor accelerates and/or aggravates the pro-fibrotic and pro-inflammatory effects of Ang II. In aim 4 we hypothesize that some of the anti-fibrotic and anti-inflammatory effects of ACEi are mediated by an increase in local and/or circulating Ac-SDKP. We will study whether a specific POP inhibitor prevents the beneficial effects of ACEi in Ang ll-induced hypertension and DOCA/salt hypertension.